Immune cells surveil aberrantly sialylated O-glycans on megakaryocytes to regulate platelet count.

Immune thrombocytopenia (ITP) is a platelet disorder. Pediatric and adult ITP have been associated with sialic acid alterations, but the pathophysiology of ITP remains elusive, and ITP is often a diagnosis of exclusion. Our analysis of pediatric ITP plasma samples showed increased anti-Thomsen-Friedenreich antigen (TF antigen) antibody representation, suggesting increased exposure of the typically sialylated and cryptic TF antigen in these patients. The O-glycan sialyltransferase St3gal1 adds sialic acid specifically on the TF antigen. To understand if TF antigen exposure associates with thrombocytopenia, we generated a mouse model with targeted deletion of St3gal1 in megakaryocytes (MK) (St3gal1MK-/-). TF antigen exposure was restricted to MKs and resulted in thrombocytopenia. Deletion of Jak3 in St3gal1MK-/- mice normalized platelet counts implicating involvement of immune cells. Interferon-producing Siglec H-positive bone marrow (BM) immune cells engaged with O-glycan sialic acid moieties to regulate type I interferon secretion and platelet release (thrombopoiesis), as evidenced by partially normalized platelet count following inhibition of interferon and Siglec H receptors. Single-cell RNA-sequencing determined that TF antigen exposure by MKs primed St3gal1MK-/- BM immune cells to release type I interferon. Single-cell RNA-sequencing further revealed a new population of immune cells with a plasmacytoid dendritic cell-like signature and concomitant upregulation of the immunoglobulin rearrangement gene transcripts Igkc and Ighm, suggesting additional immune regulatory mechanisms. Thus, aberrant TF antigen moieties, often found in pathological conditions, regulate immune cells and thrombopoiesis in the BM, leading to reduced platelet count.

Pediatric autoimmune myelofibrosis: Experience from a large pediatric tertiary care center.

Autoimmune myelofibrosis (AIMF) is a rare disorder characterized by cytopenias and autoimmunity, with characteristic bone marrow findings that include lymphocytic infiltration and fibrosis. AIMF is described predominantly in adult populations who have systemic lupus erythematosis (SLE), with scant pediatric cases described mainly in older adolescents with SLE. Here, we described the largest single-center pediatric experience of pediatric autoimmune myelofibrosis (PAIMF) series, demonstrating both similarities and distinctions from the adult experience. Patients overall respond well to steroid therapy, but these patients were significantly younger, infrequently carried a diagnosis of SLE, and causative genetic lesions were identified in many cases.

Refractory autoimmune cytopenias in pediatric Evans syndrome with underlying systemic immune dysregulation.

Evans syndrome is a rare but challenging disorder in children; and despite rapidly growing evidence for targetable systemic immune dysregulation driving these "idiopathic" autoimmune cytopenias, precision diagnosis and management remains sub-optimal among these patients. We analyzed retrospective clinical data for 60 pediatric ES patients followed at 3 large tertiary referral centers in the United States over a recent 6-year period and found that definable underlying systemic immune dysregulation was identified in only 42% of these patients throughout the course of clinical care. Median time from ES diagnosis to identification of the underlying systemic immune dysregulation disorder was 1.3 years (<1 month for rheumatologic disease, 2.3 years for CVID, 3.4 years for ALPS, and 7.4 years for monogenic disorders of immune regulation). Notably, a significantly higher percentage of patients in whom a definitive immune dysregulation disorder was ultimately identified required ≥3 cytopenia-directed therapies (92%) and also second- and third-line immunomodulatory agents (84%), vs those in whom no unifying immune dysregulation was diagnosed (65%, and 35%, respectively)-indicating that autoimmune cytopenias as a manifestation of systemic immune dysregulation are more treatment-refractory and severe. These data underline the importance of identifying the underlying systemic immune dysregulation and providing targeted therapy in pediatric ES.

COVID-19 outcomes in a large pediatric hematology-oncology center in Houston, Texas.

An understanding of the behavior of SARS-CoV-2 in pediatric hematology-oncology patients is essential to the optimal management of these patients during the COVID-19 pandemic. This study describes the characteristics and outcomes of COVID-19 disease in children with cancer or hematologic disorders treated at a large children's hospital. A retrospective cohort study was conducted at Texas Children's Cancer and Hematology Center from January 1, 2020 to September 30, 2020. All patients with a primary hematology-oncology diagnosis and SARS-CoV-2 positivity by reverse transcription polymerase chain reaction were identified. Clinical and laboratory data were obtained from the medical record. Descriptive analyses were performed to evaluate COVID-19-related outcomes and risk factors for severe disease in this population. We identified 109 patients with COVID-19 disease, including 52 hematology, 51 oncology, and 6 HSCT patients; median age was 10.3 years (IQR 4.4-15.9), and 58.7% were male. Seventy-four percent of the patients were managed in the outpatient setting. Patients with sickle cell disease were more likely to require hospitalization. ICU care was needed in 8% (n = 9) of the entire cohort, and mechanical ventilation was required in 6.4% (6 oncology patients, 1 hematology patient). COVID-19 contributed to the deaths of two cancer patients. No deaths occurred in hematology or HSCT patients. In conclusion, the risk of severe COVID-19 complications is slightly higher in pediatric hematology-oncology patients than in the general pediatric population but lower than initially feared. For most asymptomatic patients, primary disease management may continue as planned, but treatment decisions must be individualized.

Genetic variants in toll-like receptor 4 are associated with lack of steroid-responsiveness in pediatric ITP patients.

Although the most common front-line therapies for immune thrombocytopenia (ITP) have been in use for decades, it is still not possible to predict an individual patient's clinical course and response to therapy. Patients are managed with a trial-and-error approach and often suffer side effects of therapies which could have been avoided if response prediction were possible. Corticosteroids are the most frequently used upfront therapy for adults and children with ITP. Our group performed whole exome sequencing on a cohort of pediatric ITP patients, and identified two missense single nucleotide variants (SNV) in Toll-like receptor 4 (TLR4). These coding variants in TLR4 had an increased frequency in Caucasian patients with poor response to upfront steroid therapy. Both TLR4 (D299G; rs4986790) and TLR4 (T399I; rs4986791) had a minor allele frequency (MAF) of 20.7% in those patients unresponsive to steroids, but were present at lower allele frequencies of 2.3% and 3.4% in responders respectively (P < .001). These findings were consistent with the trend identified in an independent cohort of pediatric ITP patients treated with corticosteroids who underwent direct genotyping for both SNVs. This study identified two candidate genetic variants in two unique cohorts of ITP patients which may contribute to steroid response and have prognostic implications for treatment response in ITP.

Common variable immunodeficiency-associated endotoxemia promotes early commitment to the T follicular lineage.

BACKGROUND: Although chiefly a B-lymphocyte disorder, several research groups have identified common variable immunodeficiency (CVID) subjects with numeric and/or functional TH cell alterations. The causes, interrelationships, and consequences of CVID-associated CD4+ T-cell derangements to hypogammaglobulinemia, autoantibody production, or both remain unclear. OBJECTIVE: We sought to determine how circulating CD4+ T cells are altered in CVID subjects with autoimmune cytopenias (AICs; CVID+AIC) and the causes of these derangements. METHODS: Using hypothesis-generating, high-dimensional single-cell analyses, we created comprehensive phenotypic maps of circulating CD4+ T cells. Differences between subject groups were confirmed in a large and genetically diverse cohort of CVID subjects (n = 69) by using flow cytometry, transcriptional profiling, multiplex cytokine/chemokine detection, and a suite of in vitro functional assays measuring naive T-cell differentiation, B-cell/T-cell cocultures, and regulatory T-cell suppression. RESULTS: Although CD4+ TH cell profiles from healthy donors and CVID subjects without AICs were virtually indistinguishable, T cells from CVID+AIC subjects exhibited follicular features as early as thymic egress. Follicular skewing correlated with IgA deficiency-associated endotoxemia and endotoxin-induced expression of activin A and inducible T-cell costimulator ligand. The resulting enlarged circulating follicular helper T-cell population from CVID+AIC subjects provided efficient help to receptive healthy donor B cells but not unresponsive CVID B cells. Despite this, circulating follicular helper T cells from CVID+AIC subjects exhibited aberrant transcriptional profiles and altered chemokine/cytokine receptor expression patterns that interfered with regulatory T-cell suppression assays and were associated with autoantibody production. CONCLUSIONS: Endotoxemia is associated with early commitment to the follicular T-cell lineage in IgA-deficient CVID subjects, particularly those with AICs.

Association of a positive direct antiglobulin test with chronic immune thrombocytopenia and use of second line therapies in children: A multi-institutional review.

Immune thrombocytopenia (ITP) is the most common autoimmune cytopenia in children. Approximately, 25% of patients develop chronic disease, which may be unpredictable and challenging to treat. It is not currently possible to predict at the time of presentation which patients will have chronic disease or will experience symptoms requiring second-line therapy defined as treatment beyond corticosteroids, intravenous immunoglobulin, or Rh immune globulin. A multi-institutional retrospective review of 311 pediatric patients with ITP was performed with the goal of identifying clinical characteristics associated with disease course. In a cohort of 216 patients tested and for whom disease status was known, a positive direct antiglobulin test (DAT) was associated with chronic ITP vs spontaneous resolution of disease (29.2% vs 8.1%, P < 0.001) as well as the need for treatment with second line agents (38.5% vs 11.4%, P < 0.001) in 241 patients. Controlling for the effect of Evans syndrome, defined as having two immune cytopenias, a positive DAT was independently associated with chronic ITP (OR = 2.7, 95% CI: 1.0-7.2, P = 0.041) and use of second-line agents (OR: 3.6, 95% CI: 1.7-7.7, P = 0.001) by multivariate logistic regression model. These findings demonstrate an association with positive DAT and chronic disease, as well as refractory disease requiring second-line agents.

Detection of novel duplication variant in ADAMTS13 gene using chromosomal microarray analysis.

We present a case of a child with congenital thrombotic thrombocytopenic purpura found to have a compound heterozygous variant in the ADAMTS13 gene with a novel variant resulting in a large duplication of exons 9-11 of ADAMTS13 This variant was identified through additional molecular testing via a chromosomal microarray analysis. To our knowledge, this assay had not previously been utilised to identify an ADAMTS13 variant and the additional testing was possible through the involvement of a genetic counsellor.

Radiographic Union Assessment in Surgically Treated Distal Femur Fractures: A Systematic Review.

BACKGROUND: Distal femur fractures are known to have challenging nonunion rates. Despite various available treatment methods aimed to improve union, optimal interventions are yet to be determined. Importantly, there remains no standard agreement on what defines radiographic union. Although various proposed criteria of defining radiographic union exist in the literature, there is no clear consensus on which criteria provide the most precise measurement. The use of inconsistent measures of fracture healing between studies can be problematic and limits their generalizability. Therefore, this systematic review aims to identify how fracture union is defined based on radiographic parameters for surgically treated distal femur fractures in current literature. METHODS: In accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines, Medline, EMBASE, Cochrane Central Register of Controlled Trials, and Web of Science Core Collection databases were searched from inception to October 2022. Studies that addressed surgically treated distal femur fractures with reported radiographic union assessment were included. Outcomes extracted included radiographic definition of union; any testing of validity, reliability, or responsiveness; reported union rate; reported time to fracture union; and any functional outcomes correlated with radiographic union. RESULTS: Sixty articles with 3,050 operatively treated distal femur fractures were included. Operative interventions included lateral locked plate (42 studies), intramedullary nail (15 studies), dynamic condylar screw or blade plate (7 studies), dual plate or plate and nail construct (5 studies), distal anterior-posterior/posterior-anterior screws (1 study), and external fixation with a circular frame (1 study). The range of mean follow-up time reported was 4.3 to 44 months. The most common definitions of fracture union included "bridging or callus formation across 3 of 4 cortices" in 26 (43%) studies, "bony bridging of cortices" in 21 (35%) studies, and "complete bridging of cortices" in 9 (15%) studies. Two studies included additional assessment of radiographic union using the Radiographic Union Scale in Tibial fracture (RUST) or modified Radiographic Union Scale in Tibial fracture (mRUST) scores. One study included description of validity, and the other study included reliability testing. The reported mean union rate of distal femur fractures was 89% (range 58%-100%). The mean time to fracture union was documented in 49 studies and found to be 18 weeks (range 12-36 weeks) in 2,441 cases. No studies reported correlations between functional outcomes and radiographic parameters. CONCLUSION: The current literature evaluating surgically treated distal femur fractures lacks consistent definition of radiographic fracture union, and the appropriate time point to make this judgement is unclear. To advance surgical optimization, it is necessary that future research uses validated, reliable, and continuous measures of radiographic bone healing and correlation with functional outcomes. LEVEL OF EVIDENCE: Level IV. See Instructions for Authors for a complete description of levels of evidence.

Disrupted mitochondrial response to nutrients is a presymptomatic event in the cortex of the APP SAA knock-in mouse model of Alzheimer's disease.

Introduction: Reduced brain energy metabolism, mTOR dysregulation, and extracellular amyloid-ß oligomer (xcAßO) buildup characterize AD; how they collectively promote neurodegeneration is poorly understood. We previously reported that xcAßOs inhibit N utrient-induced M itochondrial A ctivity (NiMA) in cultured neurons. We now report NiMA disruption in vivo . Methods: Brain energy metabolism and oxygen consumption were recorded in APP SAA/+ mice using two-photon fluorescence lifetime imaging and multiparametric photoacoustic microscopy. Results: NiMA is inhibited in APP SAA/+ mice before other defects are detected in these amyloid-ß-producing animals that do not overexpress APP or contain foreign DNA inserts into genomic DNA. GSK3ß signals through mTORC1 to regulate NiMA independently of mitochondrial biogenesis. Inhibition of GSK3ß with lithium or TWS119 stimulates NiMA in cultured human neurons, and mitochondrial activity and oxygen consumption in APP SAA mice. Conclusion: NiMA disruption in vivo occurs before histopathological changes and cognitive decline in APP SAA mice, and may represent an early stage in human AD.

What's in a name: defining pediatric refractory ITP.

There are no agreed upon terminology to define "refractory" pediatric Immune Thrombocytopenia (ITP). Guidelines are therefore limited to arbitrary and outdated definitions. The Pediatric ITP Consortium of North America held a meeting in 2023 to define this entity. With 100% agreement, the faculty established that pediatric ITP that is refractory to emergent therapy could be defined as no platelet response after treatment with all eligible emergent pharmacotherapies. With 100% agreement, the working group established that pediatric patients with ITP that continue to demonstrate high disease burden and/or no platelet response despite treatment with multiple classes of disease modifying therapies represent a challenging subset of ITP. These patients are at higher risk of ongoing disease burden and merit additional investigation as well as consideration for clinical trials or novel therapies. Future efforts to define disease burden and disease response will be completed in collaboration with the ITP International Working Group.

Adolescent acquired thrombotic thrombocytopenic purpura: An analysis of the Pediatric Health Information System database.

The outcomes and characteristics of acquired thrombotic thrombocytopenic purpura (TTP) in adolescents is poorly understood due to an absence of studies focused on this population. To better understand the life-threatening disorder in this age, we performed an analysis of adolescent patients (ages 10-21) with TTP in the Pediatric Health Information Systems database from 2009 to 2020. The primary outcomes evaluated were in-hospital mortality and rate of TTP relapse. Secondary outcomes included rates of hemorrhagic and thrombotic complications during hospitalizations for TTP. Patients were included if they had a thrombotic microangiopathy diagnostic code, ADAMTS13 lab obtained, and received therapeutic plasmapheresis. Patients that received treatment for other non-TTP microangiopathies were excluded. A total of 99 patients with 123 hospitalizations for TTP treatment were identified. In-patient mortality occurred in 6 % (n = 6) and TTP relapse in 20 % (n = 20) of the cohort. Median time from initial admission to relapse was 33 days (IQR 15, 92). A hemorrhagic complication was identified in 29 % (n = 36) and thrombotic complication in 15 % (n = 19) of the cohort. The presence of underlying comorbidities was not associated with TTP relapse and only a diagnosis of cancer was associated with increased mortality. The rate of mortality and relapse in adolescent TTP is lower than that seen in adult registries. Long term prospective studies are needed to understand the long-term consequences of adolescent onset acquired TTP.

Risk factors and prognostic significance of platelet count abnormalities in children with HIV infection on antiretroviral therapy.

OBJECTIVES: To establish the incidence, risk factors and correlation with survival of thrombocytopenia and thrombocytosis (T/T) among children with HIV infection (CWH). DESIGN: A retrospective nested case control study of patients 0-18 years in five Baylor International Pediatric AIDS Initiative (BIPAI) centers in sub-Sahara Africa, 2004-2014. METHODS: Clinical and laboratory variables including complete blood counts (CBC) were extracted from the BIPAI electronic medical record system. Incident cases of T/T were identified and frequency-matched on follow-up time with controls with normal platelets. We calculated the prevalence and incidence density of T/T and used conditional logistic regression to evaluate their association with selected clinical variables. We constructed Kaplan-Meier curves and a Cox proportional hazards model to evaluate the impact of T/T on survival. RESULTS: Two thousand, one hundred and nine children were sampled. The incidence density of thrombocytopenia was 1 per 57.9 (95% confidence interval [CI] 50.3-66.8) CWH-years. Thrombocytopenia was higher in children with WHO Stage III/IV, lower in children on zidovudine, and had no association with use of lamivudine or nevirapine, CD4 + suppression, age, and nutrition status. Thrombocytopenia was independently associated with 2.2-fold higher mortality (95% CI 1.62-3.08). The incidence density of thrombocytosis was 1 per 11.4 (95% CI 10.7-12.1) CWH-years. Thrombocytosis was associated with higher CD4 + cell count, younger age, and use of lamivudine or nevirapine, and did not impact survival. CONCLUSIONS: Platelet count is a clinically valuable biomarker of HIV clinical progression and mortality. Laboratory studies are necessary to elucidate the mechanisms of T/T.

Cold AIHA and the best treatment strategies.

Cold-reactive autoimmune hemolytic anemia (AIHA) is rare among the hemolytic anemias. It results when 1 of a variety of processes causes the generation of immunoglobulin M (IgM) autoantibodies against endogenous erythrocytes, resulting in complement activation and predominantly intravascular hemolysis. Cold AIHA is typically a primary lymphoproliferative disorder with marrow B-cell clones producing pathogenic IgM. More rarely, secondary cold AIHA (cAIHA) can develop from malignancy, infection, or other autoimmune disorders. However, in children cAIHA is typically post infection, mild, and self-limited. Symptoms include a sequelae of anemia, fatigue, and acrocyanosis. The severity of disease is variable and highly dependent on the thermal binding range of the autoantibody. In adults, treatment has most commonly focused on reducing antibody production with rituximab-based regimens. The addition of cytotoxic agents to rituximab improves response rates, but at the expense of tolerability. Recent insights into the cause of cold agglutinin disease as a clonal disorder driven by complement form the basis of newer therapeutic options. While rituximab-based regimens are still the mainstay of therapy, options have now expanded to include complement-directed treatments and other B-cell-directed or plasma-cell-directed therapies.

Loss of RARRES1 function Promotes Follicular Lymphomagenesis and Inhibits B cell Differentiation in Mice.

Retinoic acid receptor responder 1 (RARRES1) is among the most commonly methylated loci in multiple cancers. RARRES1 regulates mitochondrial and fatty acid metabolism, stem cell differentiation, and survival of immortalized cell lines in vitro. Here, we created constitutive Rarres1 knockout (Rarres1-/-) mouse models to study RARRES1 function in vivo. Rarres1-/- embryonic fibroblasts regulated tubulin glutamylation, cell metabolism, and survival, recapitulating RARRES1 function in immortalized cell lines. In two mouse strains, loss of Rarres1 led to a markedly increased dose-dependent incidence of follicular lymphoma (FL). Prior to lymphoma formation, Rarres1-/- B cells have compromised activation, maturation, differentiation into antibody-secreting plasma cells, and cell cycle progression. Rarres1 ablation increased B cell survival and led to activation of the unfolded protein response (UPR) and heat shock response (HSR). Rarres1 deficiency had differential effects on cellular metabolism, with increased bioenergetic capacity in fibroblasts, and minor effects on bioenergetics and metabolism in B cells. These findings reveal that RARRES1 is a bona fide tumor suppressor in vivo and the deletion in mice promotes cell survival, and reduces B cell differentiation with B cell autonomous and non-autonomous functions.

Risk factors and prognostic significance of anemia in children with HIV infection on antiretroviral therapy.

OBJECTIVES: To establish the incidence, risk factors and prognostic effect of anemia in children living with HIV (CLWH). DESIGN: Retrospective nested case-control study of patients 0-18 years in five centers in sub-Saharan Africa, 2004-2014. METHODS: Incident cases of anemia were identified from electronic records and matched with CLWH without anemia. We calculated the incidence density of anemia and used conditional logistic regression to evaluate its association with risk factors, stratified by severity and type of anemia. We used a Cox proportional hazards model to evaluate the impact of anemia on survival. RESULTS: Two thousand, one hundred and thirty-seven children were sampled. The incidence density of anemia was 1 per 6.6 CLWH-years. Anemia was moderate in 31.8% and severe in 17.3% of anemia cases, which had 10-year mortality hazards of 3.4 and 4.5, respectively. Microcytic anemia (36% cases) was associated with 2.3-fold hazard of 10-year mortality, and with malnutrition and CD4 + suppression. Normocytic anemia (50.5% cases) was associated with 2.6-fold hazards of 10-year mortality, and with more severe malnutrition, CD4 + suppression, and WHO stage, but inversely associated with lamivudine and nevirapine therapy. Macrocytic anemia (13.5% cases) was neither associated with higher 10-year mortality nor with severe malnutrition or CD4 + suppression but was associated with WHO stage II/III and negatively associated with lamivudine therapy. CONCLUSION: This large multicountry study of CLWH found a high incidence density of anemia. Higher severity, normocytic and microcytic types of anemia were independently associated with long-term mortality. Laboratory studies are needed to decipher the mechanisms of anemia and how it impacts mortality in CLWH.

Romiplostim for the Treatment of Immune Thrombocytopenia: Spotlight on Patient Acceptability and Ease of Use.

Immune thrombocytopenia (ITP) is an immune-mediated disorder resulting in platelet destruction and subsequent thrombocytopenia. Bleeding symptoms range from mild cutaneous bleeding to life-threatening hemorrhage. Romiplostim, a peptide-antibody fusion product, is a thrombopoietin receptor agonist (TPO-RA) indicated for use in patients with ITP. Romiplostim is US Food and Drug Administration (FDA) approved in children ≥1 year of age with ITP of >6 months' duration who have had an inadequate response to first-line therapies or splenectomy. FDA approval in adults with chronic ITP was expanded in October 2019 to include adults with newly diagnosed (<3 months' duration) and persistent (3-12 months' duration) ITP who demonstrated an inadequate response to first-line therapies, including corticosteroids and immunoglobulins, or splenectomy. The newly published 2019 American Society of Hematology ITP Guidelines place TPO-RAs, including romiplostim, as second-line therapies in both children and adults. Here, we review the use of romiplostim as second-line therapy with a spotlight on health-related quality of life, ease of use, and patient preference.

Prostate cancer early detection program recruitment methods and show rates in men at high risk.

PURPOSE: Men with a family history of prostate cancer and black men are at higher risk for prostate cancer. Recruitment and retention of these men at high risk into early detection programs is challenging. We report a comprehensive analysis of recruitment methods, show rates and participant factors from the Prostate Cancer Risk Assessment Program, a prospective, longitudinal prostate cancer screening study. MATERIALS AND METHODS: Men 35 to 69 years old were eligible for recruitment if they had a family history of prostate cancer, were black or had a BRCA1/2 mutation. Recruitment methods were analyzed using standard statistical methods with respect to participant demographics and presentation to the first program appointment. RESULTS: Of 707 men recruited 64.9% presented to the initial program appointment. More men were recruited via radio than via referral or other methods (chi-square = 298.13, p <0.0001). Men recruited by radio were more likely to be black (p <0.001), less educated (p = 0.003) and not married or partnered (p = 0.007), and have no prostate cancer family history (p <0.001). Men recruited by referral had a higher income (p = 0.007) and were more likely to attend the initial program visit than those recruited by radio or other methods (chi-square = 27.08, p <0.0001). CONCLUSIONS: This comprehensive analysis shows that radio led to higher recruitment of black men with lower socioeconomic status. However, these men at high risk have a lower presentation rate for prostate cancer screening. Targeted motivational measures must be studied to improve the show rate for prostate cancer risk assessment in these men at high risk.